Molecular mechanisms of cell fate specification

我们的目的是确定Wnt信号如何调节将产生神经元的区域（神经外胚层）与那些不产生神经元的区域的模式。前神经外胚层区域在Wnt被拮抗的地方形成，而表皮外胚层在Wnt活跃的地方分化。至少有三种不同的Wnt通路，Wnt/ β -catenin， Wnt/PCP和Wnt/Ca+2参与分离这两种类型外胚层的结构域，至少有三种不同的Wnt信号调节因子，Dkk1， sFRP1/5和Dkk3最终在Wnt信号低的前神经外胚层表达。我们阐明了Wnt信号分支之间错综复杂、相互关联的一组相互作用，这些相互作用消除了普遍存在的、母体驱动的前神经外胚层调节状态，除了胚胎的大部分前细胞。首先，早期母源性信号通过Wnt/ β -catenin将其从后卵裂球中移除，并激活至少两个Wnt配体Wnt1和Wnt8的产生，该信号通过Wnt受体frizzle5 /8通过Wnt/JNK途径消除大多数前卵裂球的前神经外胚层命运，除了那些紧靠前极周围的细胞。Wnt/ β -连环蛋白和Wnt/JNK通路通过另一个Wnt受体frizzled1/2/7信号传导减慢；当卵裂过程中卵裂球分离，并通过前后和背腹（结- bmp）模式机制相互作用时，这一机制起着协调作用，以适当调节单个卵裂球的命运。fz5 /8依赖性的前神经调节状态的消除被前神经胚层中Wnt拮抗剂Dkk1的产生所阻断。有趣的是，Dkk1的表达依赖于Fz5/8，但随后负反馈抑制其表达。在大多数细胞中，Fz5/8的活性得以维持，部分是由于其自身转录的正反馈。在Dkk1存在的情况下，Fz5/8转录如何在前细胞中维持尚不清楚，但可能取决于Dkk3， Dkk3在前细胞中特异性表达，是Wnt信号传导的明显增强剂。这些研究揭示了早期模式中不同Wnt通路之间意想不到的和令人惊讶的复杂相互作用，以及Wnt/PCP和Wnt/Ca+2在调节早期外胚层细胞命运决定中的意想不到的作用。根据已报道的半足虫和头足虫的基因表达模式以及斑马鱼胚胎中分离的功能丧失研究，这种Wnt信号网络在后口胚胎中可能是保守的。

Our objective was to determine how Wnt signaling regulates the patterning of regions that will give rise to neurons (neuroectoderm) versus those that do not. The anterior neuroectoderm domain forms where Wnt is antagonized whereas epidermal ectoderm differentiates where Wnt is active. At least three different Wnt pathways, Wnt/beta-catenin, Wnt/PCP and Wnt/Ca+2, are involved in separating the domains of these two types of ectoderm and at least three different regulators of Wnt signaling, Dkk1, sFRP1/5 and Dkk3, are ultimately expressed in the anterior neuroectoderm where Wnt signaling is low. We elucidated an intricate, interconnected set of interactions among the Wnt signaling branches that eliminate the ubiquitous, maternally driven anterior neuroectoderm regulatory state from all but the anterior-most cells of the embryo. First, early maternally derived signaling through Wnt/beta-catenin removes it from posterior blastomeres and activates production of at least two Wnt ligands, Wnt1 and Wnt8, that signal through the Wnt/JNK pathway via the Wnt receptor, Frizzled 5/8, to eliminate anterior neuroectoderm fate from most anterior blastomeres, except those immediately around the anterior pole. Both Wnt/beta-catenin and Wnt/JNK pathways are slowed by signaling through another Wnt receptor, Frizzled 1/2/7; this serves a coordinating function to properly regulate fates of individual blastomeres as they separate during cleavage and interact via both anterior-posterior and dorsal-ventral (nodal-BMP) patterning mechanisms. Fz5/8-dependent elimination of the anterior neural regulatory state is blocked by production of the Wnt antagonist, Dkk1 in the anterior neurectoderm. Interestingly, Dkk1 expression depends on Fz5/8 but then negatively feeds back to inhibit its expression. In all but the anterior-most cells Fz5/8 activity is maintained, in part by a positive feedback on its own transcription. How Fz5/8 transcription is maintained in anterior cells in the presence of Dkk1 is not yet understood, but may depend on Dkk3, which is expressed specifically in anterior cells and is an apparent potentiator of Wnt signaling. These studies have uncovered a set of unexpected and surprisingly complex interactions among different Wnt pathways in early patterning as well as unexpected roles for Wnt/PCP and Wnt/Ca+2 in regulating early ectodermal cell fate decisions. This network of Wnt signaling is likely conserved among deuterostome embryos, based on reported gene expression patterns in hemichordates and cephalochordates and isolated loss-of-function studies in zebrafish embryos.